This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
SIRT4 / NAD-dependent protein lipoamidase sirtuin-4, mitochondrial · IHC design guide
Design Immunohistochemistry for SIRT4
Plan SIRT4 paraffin-section IHC using placental trophoblasts as a high-staining reference (HPA tissue IHC). Score cytoplasmic signal by cell type, with lymph-node germinal-center cells as a reported unstained comparison (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
SIRT4 Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Cytoplasmic in tissue IHC (HPA tissue IHC); mitochondrial matrix (UniProt) | |
| Staining pattern | Strong cytoplasmic staining in placental trophoblasts (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A03764-1) | |
| Positive control | Placenta+4 more · see all | |
| Negative control | Lymph node+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A03764-1); keep fixation consistent. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | mTORC1 represses SIRT4 expression (UniProt) | |
| Isoform / epitope | No listed isoforms; residues 1–28 precede the mature chain (UniProt) |
Section 1
Recommended SIRT4 IHC & IF Protocols
The catalog antibody protocol is paired with 4 published SIRT4 IHC protocols covering bladder, prostate, colorectal, and neuroblastoma tissue (PMC10257844; PMC9288510; PMC4522635; PMC6234993).
| Sample | Paraffin-embedded human ovary cancer tissue; fixative not specified (datasheet A03764-1) |
| Fixation | Image fixative and duration unreported (datasheet A03764-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet A03764-1) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A03764-1) |
| Primary antibody | Rabbit anti-SIRT4, 1μg/ml (datasheet A03764-1) |
| Primary incubation | Overnight at 4 °C (datasheet A03764-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A03764-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | SIRT4-positive staining in trophoblastic cells of placenta (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several cell types, most abundant in placenta. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet: A03764-1); all published protocols that specify retrieval also use citrate at pH 6 (PMC10257844; PMC9288510; PMC4522635).
Section 2
What Is the Expected SIRT4 Staining Pattern?
SIRT4 resides in the mitochondrial matrix and has no transmembrane segment (UniProt Q9Y6E7). In paraffin IHC, expect cytoplasmic staining, strongest in placental trophoblastic cells and round or early spermatids (HPA: High; cytoplasmic profile). HPA rates its tissue IHC evidence Enhanced, with medium consistency between staining and RNA data (HPA: tissue IHC).
| Cytoplasmic stain in placental trophoblastic cells or round or early spermatids. | This matches HPA's High staining in those cells and its cytoplasmic profile (HPA: tissue IHC). Mitochondrial matrix residence supports a cytoplasmic location, though chromogenic IHC may not resolve individual mitochondria (UniProt Q9Y6E7; general IHC practice). |
| Predominantly nuclear or surface outlining stain, with little cytoplasmic signal. | This conflicts with the expected mitochondrial matrix location and HPA cytoplasmic profile (UniProt Q9Y6E7; HPA: tissue IHC). Treat the pattern as suspect and review controls and morphology before assigning it to SIRT4 (general IHC practice). |
| Strong stain in an HPA-listed undetected cell population, such as lymph-node germinal center cells. | HPA reports SIRT4 not detected in those cells (HPA: tissue IHC). Assess antibody specificity or endogenous chromogen activity before interpreting the signal; a single discordant site does not prove cross-reactivity (general IHC practice). |
| Diffuse color across cells, stroma, or the whole section, obscuring cell boundaries. | This is difficult to reconcile with HPA's cell-dependent cytoplasmic pattern (HPA: tissue IHC). Background from detection reagents or inadequate washing is possible; compare a negative reagent control before scoring (general IHC practice). |
| No stain in placental trophoblastic cells or round or early spermatids. | These are HPA High reference populations, so their absence raises a run-performance or specimen concern (HPA: tissue IHC). Check section integrity and controls before concluding the sample lacks SIRT4 (general IHC practice). |
💡Expected SIRT4 appearanceA convincing positive is cytoplasmic staining in trophoblastic cells or round or early spermatids, where HPA reports High signal; dominant nuclear or surface staining is suspect given mitochondrial matrix localisation (HPA: tissue IHC; UniProt Q9Y6E7).
| Cell type and reference tissue | Use placental trophoblastic cells or round or early spermatids as high-staining references (HPA: tissue IHC). Medium glandular staining in appendix, breast, cervix, and colon offers a less intense comparison (HPA: tissue IHC). Compare the named cells, not every cell in the tissue. |
| Subcellular resolution | SIRT4 is a mitochondrial matrix protein without a transmembrane segment (UniProt Q9Y6E7). HPA describes tissue staining as cytoplasmic (HPA: tissue IHC). A resolved mitochondrial pattern can support localisation, but routine chromogenic IHC need not show distinct puncta (general IHC practice). |
| Protein processing and epitope choice | UniProt annotates a mature chain at residues 29–314 and a sirtuin-type domain at 37–314 (UniProt Q9Y6E7). If an antibody epitope is known, check whether it lies in the retained chain; no epitope is supplied here, so no antibody-specific effect can be predicted. |
| Antibody evidence and limits | HPA lists HPA029692 as IHC Enhanced and HPA029691 as IHC Approved (HPA: antibodies). The tissue profile's Enhanced rating has medium staining-to-RNA consistency (HPA: tissue IHC); neither rating guarantees specificity in a new specimen or detection setup (general IHC practice). |
| IF/ICC expectation? | UniProt's mitochondrial matrix annotation suggests intracellular, potentially mitochondrial fluorescence after appropriate cellular access (UniProt Q9Y6E7; general IF practice). HPA supplies no main IF location or ICC-IF images for SIRT4, so that appearance remains a prediction rather than an HPA-observed IF pattern (HPA: subcellular). |
| Situation | Likely cause | Next action |
|---|---|---|
| A high-reference tissue has no detectable stain. | A failed detection run, unsuitable antibody conditions, or compromised section is possible (general IHC practice); HPA reports High staining in trophoblastic cells and round or early spermatids (HPA: tissue IHC). | Inspect the named cell population and run controls, then verify the catalog antibody's IHC-P instructions and detection reagents before interpreting a biological negative (general IHC practice). |
| Only nuclei or cell borders stain strongly. | The compartment conflicts with mitochondrial matrix localisation and the HPA cytoplasmic tissue profile (UniProt Q9Y6E7; HPA: tissue IHC). | Review morphology and reagent controls; repeat with an independently validated IHC antibody if available before accepting the compartment assignment (general IHC practice; HPA: antibodies). |
| A purported negative population stains strongly. | Cross-reactivity or endogenous detection activity is possible (general IHC practice). HPA lists germinal center cells, follicle cells, and skin fibroblasts as not detected, but its survey does not establish universal biological absence (HPA: tissue IHC). | Check the exact cell type against HPA, assess a negative reagent control, and compare staining with a high-reference population in the same run (HPA: tissue IHC; general IHC practice). |
| The section shows widespread haze or precipitate. | Non-specific detection, insufficient washing, or deposited chromogen can obscure the cell-level pattern (general IHC practice). HPA's reported pattern is cytoplasmic in several cell types (HPA: tissue IHC). | Examine a negative reagent control, inspect chromogen deposits, and adjust general blocking, washing, or detection conditions as indicated by the controls (general IHC practice). |
| A medium or low reference looks weaker than placenta. | Different intensity is expected: HPA reports High trophoblastic staining, Medium glandular staining in several tissues, and Low signal in selected cells (HPA: tissue IHC). | Score the specified cells against tissue-appropriate expectations and the run's positive control; avoid requiring placental intensity in every tissue (HPA: tissue IHC; general IHC practice). |
| The proposed IF pattern is used to validate the IHC result. | HPA supplies no SIRT4 ICC-IF images or main IF location; its cytoplasmic tissue profile comes from IHC (HPA: subcellular; HPA: tissue IHC). | Treat mitochondrial fluorescence as a localisation hypothesis from UniProt, and judge the paraffin IHC slide against HPA's cell-specific tissue pattern and IHC controls (UniProt Q9Y6E7; HPA: tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing SIRT4 staining
A03764-1 · IHCIHC analysis of SIRT4 using anti-SIRT4 antibody (A03764-1). SIRT4 was detected in paraffin-embedded section of human ovary cancer tissue. He
A03764-1 · IHCIHC analysis of SIRT4 using anti-SIRT4 antibody (A03764-1). SIRT4 was detected in paraffin-embedded section of human placenta tissue. Heat m
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000089163-SIRT4/tissue/cerebral+cortex#imid_7894488
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000089163-SIRT4/tissue/colon#imid_7894430
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000089163-SIRT4/tissue/liver#imid_7894415
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000089163-SIRT4/tissue/kidney#imid_7894416
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for SIRT4 IHC & IF
🧪Run placenta first: trophoblastic cells should stain (HPA: High in trophoblastic cells). Use lymph node germinal center cells as the negative tissue (HPA: Not detected); on the placenta slide, candidate internal negative cells should show counterstain without DAB deposit, but do not designate a cell type as SIRT4-negative without validation (standard IHC control practice).
Positive control tissue: Placenta (Trophoblastic cells, HPA High)
Negative control tissue: Lymph node (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for SIRT4; derive a cell-line control from the positive tissue's cell type (Trophoblastic cells) and confirm it by RNA or western blot first.
Technical controls: Include no-primary (secondary-only) and rabbit IgG controls matched to the primary antibody’s clonality and concentration, plus a SIRT4-knockout biological negative where available (caption: rabbit anti-SIRT4; standard IHC control practice). Check placenta sections for endogenous peroxidase and biotin background before interpreting DAB signal from the biotin–SABC detection system (caption: biotinylated secondary, SABC and DAB; standard IHC control practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A03764-1 tissue-IHC caption does not state the fixative (caption: fixative unreported). That paraffin-section example used citrate retrieval at pH 6 for 20 minutes, so assess retrieval against the controls rather than treating it as a proven requirement for all specimens (caption: citrate retrieval, pH 6, 20 minutes). The supplied evidence does not establish that frozen sections or IF are easier; the HPA subcellular payload lists no ICC-IF image-bearing cell lines, and placental endogenous biotin or peroxidase could complicate interpretation with the caption’s biotin–SABC/DAB method (HPA: no ICC-IF image-bearing cell lines; caption: biotin–SABC/DAB; standard IHC control practice).
HPA tissue IHC evidence for SIRT4
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Placenta | Trophoblastic cells | High | Protein (IHC) | HPA → |
| Testis | Round or early spermatids | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
| Ovary | Follicle cells | Not detected | Protein (IHC) | HPA → |
| Skin | Fibroblasts | Not detected | Protein (IHC) | HPA → |
| Soft tissue | Chondrocytes | Not detected | Protein (IHC) | HPA → |
| Spleen | Cells in red pulp | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced SIRT4 IHC Tips
Troubleshoot SIRT4 staining in paraffin section IHC using the catalog antibody’s tissue example, reported expression patterns, and its mitochondrial matrix localisation.
How should I adjust retrieval if SIRT4 staining is weak or patchy?
Start with heat mediated retrieval in citrate buffer at pH 6 for 20 minutes (datasheet A03764-1). The catalog antibody produced a chromogenic image from a paraffin section of human ovary cancer using those conditions, although the caption does not state the fixative (datasheet A03764-1). If staining is weak, compare a modestly longer retrieval with the original condition on adjacent sections, keeping antibody concentration and development time constant (general IHC practice). Check whether stronger retrieval improves cellular signal without increasing tissue damage or diffuse background, and include the same positive control in each run (general IHC practice).
Could fixation explain absent SIRT4 staining in my paraffin sections?
Target specific fixation sensitivity is unknown: the selected tissue caption identifies a paraffin section but does not report its fixative (datasheet A03764-1). Record fixative, fixation duration, processing, section age, and retrieval conditions for each specimen before comparing staining intensity (general IHC practice). Run a known staining control alongside the problem section using citrate retrieval at pH 6 for 20 minutes, the documented condition for this antibody (datasheet A03764-1). If only the problem section fails, compare sections with known processing histories and inspect morphology; the result alone cannot identify fixation as the cause (general IHC practice).
Where should convincing SIRT4 signal appear in chromogenic IHC?
Expect predominantly cytoplasmic staining at light microscope resolution because SIRT4 resides in the mitochondrial matrix (UniProt Q9Y6E7); tissue IHC also describes cytoplasmic expression across several cell types (HPA tissue IHC). A granular cytoplasmic pattern can support that assignment, but DAB alone cannot establish mitochondrial colocalisation (general IHC practice). Mark the relevant cells on a counterstained section and compare their staining with the corresponding no primary control before assigning faint puncta (general IHC practice). Strong isolated nuclear or membranous signal warrants a specificity check because the record places SIRT4 in the matrix and lists no transmembrane segment (UniProt Q9Y6E7).
How can I assess whether unexpected staining reflects epitope accessibility or specificity?
The supplied record lists a processed SIRT4 chain spanning residues 29–314, a sirtuin type domain spanning 37–314, and no listed isoforms (UniProt Q9Y6E7). It does not locate this antibody’s epitope, so staining cannot be assigned to the processed chain solely from the image (datasheet A03764-1; UniProt Q9Y6E7). Compare retrieval conditions on matched sections while holding primary incubation and DAB development constant, then judge both staining and morphology (general IHC practice). Where suitable material is available, use an independently validated SIRT4 reagent or target depletion as an orthogonal specificity check; a no primary control addresses detection background only (general IHC practice).
How should I design IF to test the cellular identity of SIRT4 staining?
Treat IF as a separate assay: the supplied antibody example documents chromogenic staining in a paraffin section, not IF performance (datasheet A03764-1). Multiplex SIRT4 with a marker for the expected cell population; for example, trophoblastic cells show high tissue IHC staining in placenta (HPA tissue IHC). Choose spectrally separated fluorophores and place the weaker signal in a channel with lower measured tissue autofluorescence, using single stain controls to assess bleed through (general IF practice). Because SIRT4 is assigned to the mitochondrial matrix, optimise permeabilisation for access through cellular and mitochondrial membranes, then assess signal retention and morphology (UniProt Q9Y6E7; general IF practice).
What should I check when DAB background obscures SIRT4 staining?
The documented workflow used 10% goat serum block, 1 µg/mL primary antibody overnight at 4°C, a biotinylated secondary, and streptavidin biotin complex with DAB (datasheet A03764-1). Include no primary and detection reagent controls to separate antibody associated staining from detection background (general IHC practice). Check peroxidase blocking and, with this biotin based system, assess endogenous biotin interference before increasing primary antibody concentration (general IHC practice; datasheet A03764-1). Titrate primary concentration and DAB development on matched sections while checking that cellular signal remains distinguishable from stromal, edge, and other diffuse staining (general IHC practice).
How should I quantify heterogeneous SIRT4 staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and regions of interest before scoring, and apply the same inclusion rules to every section (general IHC practice). Report the percentage of positive cells and an H-score on a 0–300 scale from staining intensity and positive cell fractions, or report positive cell density per mm² for spatial comparisons (general IHC practice). Normalise cell based scores to the number of evaluable cells and area based counts to evaluable tissue area, excluding folds and necrosis consistently (general IHC practice). Keep retrieval, antibody incubation, imaging, and DAB development comparable across samples; the catalog example used 1 µg/mL primary antibody (general IHC practice; datasheet A03764-1).
How do I distinguish a credible SIRT4 positive result from artefact?
A credible result has cell associated cytoplasmic staining consistent with a mitochondrial matrix protein and a low signal in the no primary control (UniProt Q9Y6E7; general IHC practice). Use cell identity carefully: high staining is reported in placental trophoblastic cells, whereas ovarian follicle cells are listed as not detected (HPA tissue IHC). The catalog image instead shows human ovary cancer tissue, so it does not establish staining in normal follicle cells (datasheet A03764-1; HPA tissue IHC). Investigate staining confined to section edges or necrotic areas, isolated nuclear signal, and signal retained in detection controls; these patterns can indicate artefact or endogenous enzyme activity (general IHC practice; UniProt Q9Y6E7).
Boster reagents
Best SIRT4 / NAD-dependent protein lipoamidase sirtuin-4, mitochondrial IHC Antibodies
One anti-SIRT4 antibody has IHC images from human paraffin sections (A03764-1 IHC captions); the catalog lists Human, Mouse and Rat reactivity (A03764-1 catalog).
Real IHC data
Anti-SIRT4 Antibody ®
Cat # A03764-1
A03764-1 is the only SKU listed for IHC, with images from human ovary cancer, placenta, gastric cancer and glioma paraffin sections (A03764-1 applications and IHC captions). Its listed reactivity is Human, Mouse and Rat; no IF images are supplied (A03764-1 catalog).
Which to pick: For tissue IHC, choose A03764-1: its captions document staining in human paraffin sections, but do not report the fixative (A03764-1 IHC captions). For IF/ICC, there is no supported SKU to recommend here because A03764-1 does not list IF/ICC or provide IF images (A03764-1 applications and image catalog). For mouse or rat IHC, A03764-1 lists those species and IHC, although its illustrated IHC examples are human only (A03764-1 reactivity, applications and IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9Y6E7 (SIR4_HUMAN, NAD-dependent protein lipoamidase sirtuin-4, mitochondrial).
- Human Protein Atlas. SIRT4 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. SIRT4 subcellular location (ICC-IF): Highest expression in Rh30: 3.8 nTPM.
- Human Protein Atlas. SIRT4 antibody validation summary (2 antibodies).
- SIRT4 is an independent prognostic factor in bladder cancer and inhibits bladder cancer growth by suppressing autophagy. Cell division 2023 — PMC10257844.
- SIRT4 functions as a tumor suppressor during prostate cancer by inducing apoptosis and inhibiting glutamine metabolism. Scientific reports 2022 — PMC9288510.
- Tumour-suppressive function of SIRT4 in human colorectal cancer. British journal of cancer 2015 — PMC4522635.
- Tumor-suppressive function of SIRT4 in neuroblastoma through mitochondrial damage. Cancer management and research 2018 — PMC6234993.
- PubMed PMID:10381378 — UniProt-cited evidence.
- PubMed PMID:16541075 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for SIRT4 (NAD-dependent protein lipoamidase sirtuin-4, mitochondrial). Expected localisation Cytoplasmic in tissue IHC (HPA tissue IHC); mitochondrial matrix (UniProt). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A03764-1). Validated IHC antibody: A03764-1 at 1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Placenta (Trophoblastic cells); negative Lymph node. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human ovary cancer tissue; fixative not specified (datasheet A03764-1); 2. Fixation: Image fixative and duration unreported (datasheet A03764-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: Citrate pH 6 , 20 min (datasheet A03764-1); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A03764-1); 8. Primary antibody: Rabbit anti-SIRT4, 1μg/ml (datasheet A03764-1); 9. Primary incubation: Overnight at 4 °C (datasheet A03764-1). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A03764-1); keep fixation consistent. (standard IHC practice; not target-specific); Caveat: Antibody staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: No listed isoforms; residues 1–28 precede the mature chain (UniProt); Regulation: mTORC1 represses SIRT4 expression (UniProt). Evidence: UniProt Q9Y6E7, HPA tissue IHC · Enhanced, HPA cell-line RNA · Highest expression in Rh30: 3.8 nTPM, Datasheet A03764-1, PMC10257844, PMC9288510. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/SIRT4/immunohistochemistry-ihc-guide.